This invention relates to the treatment of microbial infections caused by Mycobacterium tuberculosis, Trypanosoma spp., Candida albicans, Aspergillus spp., Cryptosporidium parvum, Giardia lamblia, Plasmodium spp., Pneumocystis carinii, Toxoplasma gondii, Fusarium solani, and Cryptococcus neoformans. 
The incidence of microbial infections (e.g., mycobacterial, fungal and protozoal infections) in the immunocompromised population has significantly increased over the past several years. In particular, Candida species, especially Candida albicans, are often significant pathogens in patients infected with human immunodeficiency virus (HIV). Another pathogen, Pneumocystis carinii, causes a form of pneumonia (PCP) that is believed to be one of the leading causes of death in patients suffering from AIDS.
Human African trypanosomiasis (HAT) has reemerged as a threat to over 60 million people. Current estimates are that between 350,000 and 450,000 people are infected.
Other severe and life-threatening microbial infections are caused by Mycobacterium tuberculosis, Aspergillus spp., Cryptosporidium parvum, Giardia lamblia, Plasmodium spp., Toxoplasma gondii, Fusarium solani, and Cryptococcus neoformans. 
The antimicrobial properties of dicationic molecules have been studied since the 1930""s. Compounds of this type have typically utilized amidine groups as the cationic moieties, and their activities against a number of pathogens including Cryptosporidium parvum, Giardia lamblia, Leishmania spp., Plasmodium spp., Pneumocystis carinii, Toxoplasma gondii, Trypanosoma spp., Candida albicans, Aspergillus spp., and Cryptococcus neoformans have been reported. See e.g., King, H. et al., Ann. Trop. Med. Parasitol. 1938, 32,177-192; Blagburn, B. L. et al., Antimicrob. Agents Chemother. 1991, 35, 1520-1523; Bell, C. A. et al., Antimicrob. Agents Chemother. 1991, 35, 1099-1107; Bell, et al., Antimicrob. Agents Chemother. 1990, 34, 1381-1386; Kirk, R. et al., Ann. Trop. Med. Parastiol. 1940, 34, 181-197; Fulton, J. D. Ann. Trop. Med. Parasitol. 1940, 34, 53-66; Ivady, V. G. et al., Monatschr. Kinderheilkd. 1958, 106, 10-14; Boykin, D. W. et al., J. Med. Chem. 1995, 38, 912-916; Boykin, D. W. et al., J. Med. Chem. 1998, 41, 124-129; Francesconi et al., J. Med. Chem. 1999, 42, 2260-2265; Lindsay, D. S. et al., Antimicrob. Agents Chemother. 1991, 35, 1914-1916; Lourie, E. M; et al., Ann. Trop. Med. Parasitol. 1939, 33, 289-304; Lourie, E. M. et al., Ann. Trop. Med. Parasitol. 1939, 33, 305-312; Das, B. P. et al., J Med. Chem. 1976, 20, 531-536; Del Poeta, M. et al., J. Antimicrob. Chemother. 1999, 44, 223-228; Del Poeta, M. et al., Antimicrob. Agents Chemother. 1998, 42, 2495-2502; Del Poeta, M. et al., Antimicrob. Agents Chemother. 1998, 42, 2503-2510.
Despite the broad range of activity exhibited by diamidines, only one compound of this chemical type, pentamidine, has seen significant clinical use. Pentamidine has been used clinically against African trypanosomiasis, antimony-resistant leishmaniasis and P. carinii pneumonia. See e.g., Apted, F. I. C., Pharmacol. Ther. 1980, 11, 391-413; Bryceson, A. D. M. et al., Trans. Roy. Soc. Trop. Med. Hyg. 1985, 79, 705-714; Hughes, W. T.; et al., Antimicrob. Agents Chemother. 1974, 5, 289-293.
A number of compounds in this class of dicationic molecules have been shown to bind to the minor-groove of DNA at AT-rich sites and the details of their interaction with the minor-groove have been elucidated from biophysical studies and from several crystal structures. It is hypothesized that these types of molecules exert their biological activity by first binding to DNA and then by inhibiting one or more of several DNA dependent enzymes (i.e., topoisomerases, nucleases, etc.) or possibly by direct inhibition of transcription. See, Tanious, F. A. et al., J. Biomol. Struct. and Dyn. 1994, 11, 1063-1083.; Wilson, W. D. et al., J. Am. Chem. Soc. 1998, 120, 10310-10321; Bailly, C. et al., Anti-Cancer Drug Design, 1999, 14, 47-60; Mazur, et al., J. Molecular Biology 2000, 300, 321-337; Trent, J. O.; et al., J. Med. Chem. 1996, 36, 4554-4562; Guerri, A. et al., Nucleic Acids Res. 1998, 26, 2873-2878; Laughton, C. A. et al., Biochemistry 1996, 35, 5655-5661; Beerman, T. A. et al., Biochim. Biophys. Acta 1992, 1131, 52-61; Bell, C. A.; et al., Antimicrob. Agents Chemother. 1993, 37, 2668-2673; Dykstra, C. C. et al., Antimicrob. Agents Chemother. 1994, 38, 1890-1898; Hildebrandt, E. et al., J. Euk. Microbial. 1998, 45, 112-121; Henderson, D. et al., Nature Medicine 1995, 1, 525-527; Fitzgerald, D. J.; et al., J. Biol. Chem 1999, 274, 27128-27138.
2,5-Diphenylfuran and 2,4-diphenylfuran diamidines have been found to be highly effective treatments in animal models for Pneumocystis carinii and Cryptosporidium parvum. See Blagburn, B. L. et al., Antimicrob. Agents Chemother. 1991, 35, 1520-1523; Boykin, D. W. et al., J. Med. Chem. 1995, 38, 912-916; Boykin, D. W. et al., J. Med. Chem. 1998, 41, 124-129; Francesconi, I. et al., J. Med. Chem. 1999, 42, 2260-2265; Tidwell, R. R. J. Parasitol. 1998, 84, 851-856. Furthermore, these molecules have shown antifungal activity in vitro against Candida albicans and Cryptococcus neoformans. See, Del Poeta, M. et al., J. Antimicrob. Chemother. 1999, 44, 223-228; Del Poeta, M. et al., Antimicrob. Agents Chemother. 1998, 42, 2495-2502; Del Poeta, M. et al., Antimicrob. Agents Chemother. 1998, 42, 2503-2510.
Although there are reports of antimicrobial activity of guanidino compounds, this class of cationic compounds has not been studied as extensively as their amidino analogs. See Lourie et al., Ann. Trop. Med. Parasitol. 1937, 31, 435-445.
The synthesis, DNA-binding affinities and antimicrobial properties of 2,5-bis{[alkyl(or aryl)imino]aminophenyl}furans and thiophenes are described. These compounds have the imino group of the amidine attached to an xe2x80x9canilinoxe2x80x9d nitrogen (in contrast to the known amidino furans in which the imino group is directly attached to the aryl ring). These compounds, hereafter, are referred to as xe2x80x9creversedxe2x80x9d amidines. The various effects of placing substituents on the central phenyl rings of the 2,5-diphenylfuran framework of this class of compounds are also described.
One aspect of the invention relates to novel compounds that are useful in treating microbial infections caused by Mycobacterium tuberculosis, Trypanosoma spp., Candida albicans, Aspergillus spp., Cryptosporidium parvum, Giardia lamblia, Plasmodium spp., Pneumocystis carinii, Toxoplasma gondii, Fusarium solani, and Cryptococcus neoformans. Compounds of the present invention have a structure according to Formula I: 
wherein:
R1, R2, R3 and R4 are each independently selected from the group consisting of H, alkyl, alkoxy, halide, and alkylhalide groups;
R5 is H, alkyl or aryl;
R6 is H, alkyl, aryl, or NR7R8, wherein R7 and R8 are each independently selected from the group consisting of H, alkyl and aryl; and
X is O, S or NR9, wherein R9 is H or alkyl.
Additional aspects of the invention include pharmaceutical compositions comprising a compound having a structure according to Formula I, or a pharmaceutical salt thereof (i.e., an xe2x80x9cactive compoundxe2x80x9d), in a pharmaceutically acceptable carrier. Pharmaceutical compositions of the present invention are useful in the treatment of microbial infections caused by Mycobacterium tuberculosis, Trypanosoma spp., Candida albicans, Aspergillus spp., Cryptosporidium parvum, Giardia lamblia, Plasmodium spp., Pneumocystis carinii, Toxoplasma gondii, Fusarium solani, and Cryptococcus neoformans. 
Certain aspects of the invention relate to methods of treating microbial infections caused by Mycobacterium tuberculosis, Trypanosoma spp., Candida albicans, Aspergillus spp., Cryptosporidium parvum, Giardia lamblia, Plasmodium spp., Pneumocystis carinii, Toxoplasma gondii, Fusarium solani, and Cryptococcus neoformans, in a subject in need of such treatment. The method comprises administering to the subject a compound according to Formulas (I), or a pharmaceutically acceptable salt thereof, in an amount effective to treat the microbial infection.
A further aspect of the present invention is the use of the active compounds described herein for the manufacture of a medicament for the treatment of microbial infections caused by Mycobacterium tuberculosis, Trypanosoma spp., Candida albicans, Aspergillus spp., Cryptosporidium parvum, Giardia lamblia, Plasmodium spp., Pneumocystis carinii, Toxoplasma gondii, Fusarium solani, and Cryptococcus neoformans in a subject in need of such treatment.
The foregoing and other aspects of the present invention are explained in detail in the specification set forth below.